Gene silencing is a core cellular regulatory mechnaism. One form of silencing, conserved across eukaryotic kingdoms, involves histone H3 lysine 9 (H3K9) methylation, proteins of the HP1 family that recognize this modification, and, intriguingly, the RNAi machinery. Despite the conservation and significance of this mechanism, the fundamental questions of how this type of silencing is initiated, how RNAi promotes histone methylation, and how repression of gene expression is ultimately effected remain largely unanswered. To address these issues, we recently shifted our efforts from studies of the histone variant H2A.Z and other molecules that regulate silencing in S. cerevisiae (which lacks both H3K9 methylation and RNAi systems) to studies of silencing in the highly tractable fission yeast Schizosaccharomyces pombe. In this proposal, we seek to capitalize on a series of preliminary investigations in fission yeast already carried out in our laboratory to achieve a mechanistic understanding of gene silencing. This work will inform our understanding of repressive histone methylation, a mechanism that has been strongly implicated in the inactivation of tumor suppressor genes in human cancers. PUBLIC HEALTH RELEVANCE: Our work focuses on understanding the mechanism of gene silencing mediated by repressive histone methylation. Since such mechanisms have been strongly implicated in the inactivation of tumor suppressor genes in human cancers, our fundamental insights into this conserved process will inform more applied studies of tumor biology. Our hope is that this will open up new avenues for the development of therapies that reactivate the expression of tumor suppressor genes and arrest growth and/or induce death of malignant cells.